Low Antithrombin Research Articles

Hämostase und Präeklampsie

A longitudinal study has been undertaken in 125 pregnant women between 25 and 40 weeks of gestation, to provide systematic information on the changes that occur in a wide range of haemostasiological and haemorheological variables. Fibrinogen, D-Dimer, Factor VIIIR: Ag, erythrocyte aggregation and plasma viscosity rose markedly throughout pregnancy. Antithrombin III and alpha 2-antiplasmin were unchanged during pregnancy. There were no significant differences between women in the pre-eclamptic group (N = 16) and the control group. HELLP syndrome (N = 7) was associated with high D-Dimer (p less than 0.05), and TAT (p less than 0.05), low antithrombin III (p less than 0.03), protein C (p less than 0.01) and platelets (p less than 0.001). Our results demonstrate that during pregnancy (also, however, in pre-eclamptic women) alterations of the coagulation system occur, but these changes do not affect the overall haemostatic balance. Findings in patients with "true" HELLP syndrome are consistent with an increased tendency for intravascular coagulation.

Antithrombin III Activity Is Not Related to Plasma Homocysteine Concentrations

Arterial and venous thromboembolic events represent frequent and life-threatening complications in homocystinuric patients and are responsible for their early deaths. Reduced levels of antithrombin III activity in homocystinuric patients have recently been reported. So, high plasma L-homocysteine concentration could play a role in the low antithrombin III activity level. In the present study, we have studied the relationship between total plasma homocysteine and inhibitors of blood coagulation levels in 16 patients with malignancies who received bone marrow grafts. There were no correlations between homocysteine values and inhibitors of blood coagulation levels. So, while the defect in amino acid transsulfuration that is responsible for homocystinuria can directly affect the synthesis or activity of some clotting factors, homocysteine concentration is not responsible for this effect.

Apheresis of low density lipoproteins using a heparin-based sorbent with low antithrombin III binding capacity

A sorbent based on heparin fraction with low affinity for antithrombin III is proposed for low density lipoproteins apheresis in hypercholesterolemia. Heparin was fractionated on antithrombin III-Sepharose; fractions with high and low affinity for antithrombin III were immobilized on CNBr-activated Sepharose 4B. Both sorbents appeared to have an LDL-binding capacity essentially similar to that of the sorbent based on unfractionated heparin. However only plasmasorption on a low affinity heparin-containing sorbent did not reduce plasma antithrombin III. Hence the use of this sorbent may be advantageous over the currently applied sorbents with unfractionated heparin in the treatment of familial hypercholesterolemia.

Predisposing factors of recurrent embolization in cardiogenic cerebral embolism.

To elucidate the pathophysiology of intracardiac thrombus formation, serial two-dimensional echocardiographic examinations were performed on 30 consecutive patients with acute cardiogenic cerebral embolism in parallel with measurement of hematocrit and plasma levels of antithrombin III. The data from groups of patients with and without newly formed or enlarged thrombi were compared. Intracardiac thrombi were detected in eight of the 30 patients (27%), four at admission and four after admission. Enlargement of the thrombus was observed in four, and systemic embolization recurred in three of the eight. Antithrombin III levels already were low at admission in patients who later developed thrombi or had enlarged thrombi on serial examinations. When the development or enlargement of an intracardiac thrombus was detected by echocardiography, the diameter of the inferior vena cava was found to be reduced. At the same time, a decrease in antithrombin III and an increase in hematocrit were demonstrated. Intracardiac thrombi are frequently detected by repeated echocardiographic examination in patients with cerebral embolism. Dehydration seems to accelerate thrombus formation that is reflected by a decrease in antithrombin III. A low antithrombin III level at admission and/or a decrease in antithrombin III after admission may indicate the possible recurrence of embolism.

Relationship Between renal Histology and Plasma Antithrombin III Activity in Women with Early Onset Preeclampsia

Renal biopsy was performed in 12 women with the clinical diagnosis of severe, early-onset preeclampsia at the time of cesarean delivery for the express purpose of aiding future counseling on the risk of recurrence. The mean gestation at delivery was 30 +/- 3 weeks. The mean birthweight was 1090 +/- 505 gm. Four women (33%) were multiparous. Antithrombin III activity was determined immediately prior to delivery unrelated to clinical care and as part of other protocols. The biopsy was performed without difficulty in each, although the sample was inadequate in one patient. The clinical diagnosis of preeclampsia was confirmed in nine (82%). However, three of the nine had underlying renal disease, as did the two women without histologic evidence of preeclampsia (42% of the total). Correlations between laboratory parameters with the histopathologic diagnoses were sought. Neither uric acid, creatinine, blood urea nitrogen, platelet count, or 24-hour urinary protein measurements aided the differentiation of the various subgroups. Antithrombin III activity in women with biopsy-supported preeclampsia (77% +/- 12%) was significantly lower than that in women without histologic evidence of preeclampsia (116% +/- 8%). Antithrombin III activity correctly predicted biopsy findings in at least 9 of 11 (82%). These preliminary findings confirm the high frequency of underlying disease in women with early-onset preeclampsia. Although low antithrombin III activity does not differentiate between "pure" preeclampsia and superimposed disease, a normal antithrombin III activity is reassuring and more consistent with a nonpreeclamptic renal complication than with preeclampsia.

Isolation of Nine Human Plasma Proteinase Inhibitors by Sequential Affinity Chromatography

Purification of nine plasma proteinase inhibitors and one zymogen from a single batch of human plasma, using affinity chromatography has been accomplished. Those isolated were plasminogen (lysine-Sepharose), alpha-2-antiplasmin (plasminogen-Sepharose), high and low molecular weight kininogens (CM-papain-Sepharose), alpha-2-macroglobulin (Zn++ chelate-Sepharose), alpha-1-proteinase inhibitor, alpha-1-antichymotrypsin, Cl-inhibitor, inter-alpha-trypsin inhibitor (Blue-Sepharose) and antithrombin III (heparin-Sepharose). Alpha-2-macroglobulin and alpha-1-proteinase inhibitor required gel filtration as additional purification steps. Each protein was recovered in both high yield and purity.

Coagulation Parameters in Maternal and Cord Blood at Delivery

A wide range of hemostatic variables were studied in 300 maternal/neonatal pairs at the time of delivery and also in 375 healthy nonpregnant women who served as controls. There was significant prol...

Influence of plasma protease activation on electroimmunoassay and nephelometry of plasma fibronectin in sepsis

In vitro experiments have shown that proteases such as trypsin, kallikrein and plasmin may split plasma fibronectin, yielding changes in apparent mass concentration, i.e.: a decrease when using immunonephelometry (IN), and an increase when using electroimmunoassay (EIA). In the present in vivo study, plasma from 49 patients with severe infections was assayed for fibronectin by IN and EIA, and for prekallikrein, plasminogen and antithrombin. In patients with normal prekallikrein (n=26) and plasminogen (n=23), the agreement between the two fibronectin assay methods was good (ratio EIA/IN = 0.99 ± 0.06); a similar good agreement was found in 45 healthy blood donors (ratio 0.97 ± 0.02). In contrast, the 20 patients with low prekallikrein showed fibronectin values that were significantly higher by EIA than by nephelometry (ratio 1.27 ± 0.10, p<0.01). Similarly, the 26 patients with low plasminogen had a significantly increased ratio (1.21 ± 0.09, p<0.05). No such difference was seen, however, between patients with low or normal antithrombin. Thus, kallikrein and plasmin activation in vivo appear to increase the fibronectin concentration measured by EIA, possibly due to the formation of small fragments with increased electrophoretic motility.

Low antithrombin III: accurate predictor of idiopathic respiratory distress syndrome in premature neonates.

A prospective study was performed in premature neonates to determine the predictive values of antithrombin III (AT III) deficiency immediately after birth, for the subsequent development of idiopathic respiratory distress syndrome (IRDS), intraventricular haemorrhage (IVH) and death. Of the 81 premature infants studied, 24 developed IRDS (30%). Of these 24 premature infants, 8 also developed IVH and 9 infants died within the follow-up period of 7 days. The mean plasma AT III level was significantly lower in the infants developing IRDS (0.23 U/ml vs 0.35 U/ml, P less than 0.0005). Within this study group 33 neonates of less than 30 weeks' gestation showed a prevalence for IRDS of 48%. In this group, AT III activity levels below 0.30 U/ml were 8.5 times as likely to result from infants with IRDS than from infants without IRDS. The diagnostic accuracy indices of criteria for the development of IRDS were: a sensitivity of 100%, a specificity of 88%, a positive predictive value of 89% and a negative predictive value of 100%. The predictive values for the development of IVH and occurrence of death were insignificant. Therefore, in premature neonates the combination of less than 30 weeks' gestation and an AT III below 0.30 U/ml is highly suggestive of IRDS and may facilitate the evaluation of early treatment.

Intraperitoneal heparin in peritoneal dialysis and its effect on fibrinopeptide A in plasma and dialysate.

In 6 patients on continuous ambulatory peritoneal dialysis we investigated the inhibition of intraperitoneal fibrin formation by heparin. A continuous addition of 500 U of heparin per liter dialysate was used for 52 h. In plasma no heparin activity could be detected, even 52 h after intraperitoneal administration of heparin. The fibrin formation was determined by fibrinopeptide A, a thrombin-induced split product of fibrinogen. In patients under regular continuous ambulatory peritoneal dialysis we determined the fibrinopeptide A concentrations in plasma. The values were comparable with the fibrinopeptide A concentrations measured in disseminated intravascular coagulopathy. They decreased during intraperitoneal administration of heparin from 63.2 +/- 11.8 to 4.9 +/- 1.7 ng/ml. The fibrinopeptide A concentration in the 4-hour intraperitoneal dialysate (155.8 +/- 15.7 ng/ml) decreased after heparin administration to 8.5 +/- 2.0 ng/ml and was always higher than in plasma. We conclude that 500 U heparin per liter dialysate prevents the intraperitoneal fibrin formation. The low antithrombin III concentration (0.44 +/- 0.13 mg/dl) in protein-poor dialysate seems to be sufficient to inhibit the thrombin activity after acceleration by heparin.

Hemodynamic and hemorheological profiles in women with proteinuric hypertension of pregnancy and in pregnant controls

We obtained blood samples from 52 patients with pre-eclampsia and from 40 pregnant controls for measurement of plasma urate levels, hematocrit, white cell count and various hemorheological parameters. We also used impedance cardiography to measure cardiac output in both groups and from the results derived values for total peripheral resistance and oxygen transport. Central venous pressure was measured with a superior vena cava catheter in patients with pre-eclampsia but not in controls. Women with pre-eclampsia had significantly lower cardiac output and central venous pressure when compared with a control group. A modest correlation was observed between central venous pressure and cardiac output. The majority of pre-eclamptic patients had significantly raised hematocrit, leucocyte count, uric acid and red cell aggregation. Red cell deformability was significantly decreased in patients with pre-eclampsia. Most patients with severe pre-eclampsia (BP diast. greater than 100 mmHg) had a low Antithrombin III and colloid osmotic pressure level. The leucocyte count was raised when compared with the women with moderate pre-eclampsia. Oxygen delivery was reduced in patients with pre-eclampsia because of impaired rheological properties of their blood.

The mechanism of reduced antithrombin III activity in women with preeclampsia.

The cause(s) of the low plasma antithrombin III activity in women with preeclampsia is unknown. The purpose of this study was to evaluate the relationship between plasma antithrombin III activity and both clotting parameters (platelet count and fibrinopeptide A) and renal parameters (serum creatinine and uric acid concentration). Forty-seven preeclamptic women were studied within the 2-day interval before delivery. Creatinine did not correlate significantly with any of the other study parameters. However, there were significant correlations between antithrombin III activity and platelet count (r = 0.53, P less than .005) and between antithrombin III and fibrinopeptide A (r = -0.29, P less than .05). The correlation between platelet count and fibrinopeptide A was not significant. These findings support the hypothesis that antithrombin III consumption by fibrin generation is initiated by platelet activation, and is at least one cause of the low activity level known to occur in women with preeclampsia.

Mapping and quantification of the major oligosaccharide components of heparin.

A new method of determining the oligosaccharide composition of commercial glycosaminoglycan heparin is described in which heparin was first depolymerized using heparin lyase (EC 4.2.2.7), and then analysed by a single h.p.l.c. step. All 20 of the porcine and bovine heparins examined were found to contain a small number of major oligosaccharide components, which on average comprised 86% of their mass. The five most abundant oligosaccharides have defined chemical structures. Although the relative abundance of oligosaccharides varied, the heparins examined were surprisingly similar. Porcine, bovine, low-Mr, and high and low antithrombin III (ATIII)-affinity heparins, however, each had distinctly different proportions of these major oligosaccharide components. The concentrations of one of these five oligosaccharides, containing a portion of the ATIII binding site, correlated with the anticoagulant activity of the ATIII-affinity-fractionated porcine-mucosal heparins from which it was derived. An additional oligosaccharide of undetermined structure was found in significant quantities in both bovine heparin and high ATIII-affinity porcine-mucosal heparin. The correlation between oligosaccharide concentration and anticoagulant activity suggests that the oligosaccharide is derived from a structural variant of the ATIII-binding site. Finally, for the heparins examined chondroitin/dermatan sulphate formed 0.6-7.4% of their mass.

Antithrombin III Utah: proline-407 to leucine mutation in a highly conserved region near the inhibitor reactive site

A dysfunctional antithrombin III (ATIII) gene encoding a qualitatively and quantitatively abnormal anticoagulant molecule is responsible for hereditary thrombosis in a Utah kindred [Bock et al. (1985) Am. J. Hum. Genet. 37, 32-41]. Nucleotide sequencing of the entire protein-encoding portion of the cloned ATIII-Utah gene revealed a C to T transitional mutation which converts proline-407 to leucine. Proline-407 is located 14 amino acids C-terminal to the reactive site arginine of ATIII in a core region of the molecule that has been highly conserved during evolution of the serine protease inhibitor (serpin) gene family. The location of this proline in the crystal structure of the homologous serpin alpha 1-antitrypsin suggests that the leucine substitution in ATIII-Utah may interfere with correct folding of the mutant gene product, leading to its rapid turnover and the low antithrombin levels observed in patient plasmas. The Pro-407 to Leu mutation does not interfere with binding of antithrombin III to heparin. Patient antithrombin III, isolated by affinity chromatography on heparin-Sepharose, was reacted with purified thrombin. ATIII encoded by the patient's normal gene formed protease-inhibitor complexes with thrombin, whereas the product of the ATIII-Utah gene did not. The Pro-407 to Leu mutation destroys a restriction site for the enzyme StuI, permitting rapid diagnosis of affected members of the Utah kindred by Southern blotting of genomic DNA.

Plasma vitronectin polymorphism in normal subjects and patients with disseminated intravascular coagulation

Vitronectin, also known as serum-spreading factor or S-protein, mediates cell adhesion and inhibits formation of the membrane-lytic complex of complement and the rapid inactivation of thrombin by antithrombin III in the presence of heparin. Vitronectin is normally present in plasma at a concentration of approximately 300 micrograms/mL. The investigators quantified plasma vitronectin with an enzyme-linked immunosorbent assay and visualized reduced and nonreduced vitronectin by immunoblotting after separation of plasma or serum by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The concentration of plasma vitronectin was markedly reduced in some patients with disseminated intravascular coagulation, especially in those with liver failure; it was near normal in patients with metastatic cancer and acute leukemia. Patients with vitronectin levels less than 40% normal invariably had low fibrinogen and antithrombin III and a prolonged prothrombin time. In both normal and patient plasmas there was heterogeneity in the ratio of the 75,000- and 65,000-mol wt polypeptides of reduced vitronectin: 18% had mostly the 75,000-mol wt polypeptide, 59% had roughly equal amounts of the two polypeptides, and 22% had mostly the 65,000-mol wt polypeptide. This polymorphism is inherited and appears to be due to two alleles that are present with approximately equal frequency. The blotting patterns of vitronectin in reduced and nonreduced plasmas were largely unaltered in plasma of patients with defibrination syndrome, fibrinolysis, liver failure, sepsis, metastatic cancer, and acute leukemia. There was no evidence of fragmentation of vitronectin or formation of the disulfide-bonded complex of vitronectin and thrombin-antithrombin III that is found when blood is clotted. Thus these results corroborate in vitro observations that the liver is the major source of plasma vitronectin, suggest that vitronectin may become depleted during disseminated intravascular coagulation, and define a genetic polymorphism of vitronectin.

Plasma Vitronectin Polymorphism in Normal Subjects and Patients With Disseminated Intravascular Coagulation

Abstract Vitronectin, also known as serum-spreading factor or S-protein, mediates cell adhesion and inhibits formation of the membrane-lytic complex of complement and the rapid inactivation of thrombin by antithrombin III in the presence of heparin. Vitronectin is normally present in plasma at a concentration of approximately 300 micrograms/mL. The investigators quantified plasma vitronectin with an enzyme-linked immunosorbent assay and visualized reduced and nonreduced vitronectin by immunoblotting after separation of plasma or serum by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The concentration of plasma vitronectin was markedly reduced in some patients with disseminated intravascular coagulation, especially in those with liver failure; it was near normal in patients with metastatic cancer and acute leukemia. Patients with vitronectin levels less than 40% normal invariably had low fibrinogen and antithrombin III and a prolonged prothrombin time. In both normal and patient plasmas there was heterogeneity in the ratio of the 75,000- and 65,000-mol wt polypeptides of reduced vitronectin: 18% had mostly the 75,000-mol wt polypeptide, 59% had roughly equal amounts of the two polypeptides, and 22% had mostly the 65,000-mol wt polypeptide. This polymorphism is inherited and appears to be due to two alleles that are present with approximately equal frequency. The blotting patterns of vitronectin in reduced and nonreduced plasmas were largely unaltered in plasma of patients with defibrination syndrome, fibrinolysis, liver failure, sepsis, metastatic cancer, and acute leukemia. There was no evidence of fragmentation of vitronectin or formation of the disulfide-bonded complex of vitronectin and thrombin-antithrombin III that is found when blood is clotted. Thus these results corroborate in vitro observations that the liver is the major source of plasma vitronectin, suggest that vitronectin may become depleted during disseminated intravascular coagulation, and define a genetic polymorphism of vitronectin.

Preeclampsia, delivery, and the hemostatic system

To determine the effects of preeclampsia and delivery, the hemostatic system was evaluated before and 24 to 48 hours after delivery in 59 nulliparous patients without clinical signs of disseminated intravascular coagulation. Fifteen patients with mild preeclampsia and 18 with severe preeclampsia were compared with 26 pregnant control patients. Preeclampsia was associated with high fibronectin (p less than 0.001), low antithrombin III (p less than 0.001), and low alpha 2-antiplasmin (p less than 0.005), suggesting endothelial injury, clotting, and fibrinolysis, respectively. After delivery, fibronectin decreased only in preeclamptic patients (p less than 0.005); alpha 2-antiplasmin increased in all groups (p less than 0.001). Endothelial injury in preeclampsia appeared to resolve soon after delivery, which could contribute to the rapid clinical improvement noted in the early puerperium.

Preeclampsia is Not Associated with Excess Fetal Clotting

Paired maternal and umbilical vein blood samples were obtained from 15 women and the plasma antithrombin III activity and fibrinopeptide A concentration measured. Women with preeclampsia had significantly lower antithrombin III activities and higher fibrinopeptide A concentrations (P less than .05) than normals. However, there were no significant differences between the two groups of fetuses. Thus, thrombin generation was not increased in the fetuses delivered from preeclamptic women.

α2-MACROGLOBULIN AND ANTITHROMBIN III ARE EQUALLY IMPORTANT INHIBITORS OF THROMBIN IN NEONATAL PLASMA

Healthy newborn infants rarely Buffer from thrombosis, despite their low plasma antithrombin III (ATIII) levels. In adults, ATIII is the main inhibitor of thrombin; α2-macroglobulin (α2M) and heparin-cofactor (HCII) are of lesser importance. In infants and children, plasma α2M concentrations are higher than in adults and exceed ATIII activities by about 2-fold. Therefore we determined the contribution of all 3 antiproteases to the inhibition of 125I-thrombin in defibrinated pooled adult and cord plasmas. Thrombin-inhibitor complexes were quantitated by SDS-PAGE. Thrombin was inhibited to a similar extent in both plasmas. The proportions of thrombin complexed to α2M, ATIII and HCII are summarized below:Values are means (SD) of 18 determinations/pooled plasma. The observed differences in distributions are consistent with calculated velocities which favour the complex formation between α2M and thrombin in neonatal plasma. Upon addition of heparin, ATIII became the most important inhibitor of thrombin in cord plasma.Conclusion: In the absence of heparin, α2M and ATIII are equally important inhibitors of thrombin in neonatal plasma. Speculation; High α2M plasma concentrations may protect healthy newborn infants and possibly also ATIII deficient children from thrombosis.

Hemostatic alterations in Crohn's disease.

SummaryThromboembolic disease is a known complication of inflammatory bowel disease, and especially of ulcerative colitis, but is less frequently reported in Crohn’s disease. In this study, we have determined 16 coagulation parameters in 20 consecutive patients with Croh n s disease at the time of admission to the hospital.The patients were divided into two subgroups of ten patients each, one with a Crohn’s Disease Activity Index (CDAI) higher than 300 and one lower than 300. No differences concerning age, sex, body mass index, lengh of the history of illness, main site of disease, and treatment scheme between both subgroups were Present. A CDAI value of higher than 300, corresponding with a more severe clinical stage, and with a higher rate of complications during the observation period was associated with a significantly higher platelet count, an increased fibrinogen level, a longer prothrombin time, and a lower antithrombin III value.